Teeth are essential organs for animal survival and of obvious clinical and/or cosmetic importance. There are many instances where tooth replacement is desirable and current treatments are restricted to artificial prostheses or implants.
Tooth primordia explants can be cultured in vitro allowing a variety of manipulation studies including introduction of genes and/or proteins and tissue recombinations.
Tooth development requires the combination of cells from mesenchymal and epithelial lineages. Development of the mammalian tooth has been recognised as a model system for study of epithelial/mesenchymal interactions during organogenesis. Teeth start to develop early in mammalian embryogenesis (11 days in mice, 6 weeks in humans), from a series of reciprocal interactions between two cell types: oral epithelial cells and neural crest-derived mesenchyme cells.
Inductive signals for tooth development come from the epithelium whereupon the responding mesenchymal cells are programmed to become odontogenic (2).
Odontogenic mesenchymal cells then provide instructive signals for further tooth development (3). The epithelial cells eventually give rise to ameloblasts which are responsible for enamel formation and mesenchyme cells form odontoblasts which produce dentine and cementoblasts.
The identity of these different instructive signals has been revealed by gene expression studies and implantation experiments. FGF8, BMP4 and SHH are established as early instructive signals from the oral epithelium (3). BMP's, FGF's and activin are among the early signals from the mesenchyme (3. 4).
Prior art approaches to the production of tooth primordia have included in vitro tissue recombination. In this approach, two different tissue types are independently dissected from the animal embryo, and these tissues are recombined in the laboratory. Signals from one may then induce formation of tooth primordia in the other. This is a labour intensive process carried out by highly trained workers involving a great deal of surgical skill.
In an alternative approach, Young et al. has showed that cells dissociated from early tooth buds when cultured on a matrix and implanted into an adult animal can form teeth, indicating the presence of both epithelial and mesenchymal dental stem cells (Young, C. S., Terada, S., Vacanti, J. P., Honda, M., Bartlett, J. D., Yelick, P. C. (2002) Tissue engineering of complex tooth structures on biodegradable polymer scaffolds. J. Dent. Res. 81, 695-700).
For human therapeutic purposes the major drawback is the potential problem of graft rejection, thus requiring either immunosuppression of host (recipient), or genetic manipulation of the graft cells to circumvent rejection, and the difficulty of obtaining the cells. Therefore the use of cells derived exclusively from each patient would avoid such rejection problems.
Sharpe (WO 01/60981) showed that cultured embryonic stem cells can give rise to epithelial and mesenchyme lineages, enabling the production of teeth primordia from embryonic stem cells.
Use of stem cells such as embryonic stem cells requires purification and expansion of a population of cells. This involves intricate and highly skilled separation and manipulation techniques. Accordingly, a method for the provision of cells to form tooth progenitor cells for implantation which does not require separation and purification, would greatly facilitate the production of such an implant.
Another problem associated with the use of embryonic stem cells is the limited availability and ease with which the embryonic stem cells can be obtained.
Adult (i.e. non-embryonic) bone marrow cells are known to contain populations of stem cells and pluripotential cells which give rise to (a) haematopoietic cells and (b) stromal (mesenchymal) cells. Haematopoietic cells in the bone marrow, however, do not give rise to non-haematopoietic tissues (Wagers et al). Mesenchymal stem cells give rise to homogeneous differentiated cell types of tissues including bone, cartilage, fat, muscle, tendon, hematopoiesis-supporting stroma, and mesangial tissues, but are not known to be capable of forming organs of composite cell lineages and those that require specific reciprocal tissue interactions, such as teeth whose development requires contributions from more than one cell lineage.
The present invention seeks to overcome at least some of the problems associated with the prior art.